Zähner D, Sener A, Malaisse W J
Laboratory of Experimental Medicine, Brussels Free University, Belgium.
Biochim Biophys Acta. 1990 Jul 12;1053(2-3):125-9. doi: 10.1016/0167-4889(90)90003-v.
A radioisotopic method for the assay of reduced or oxidized pyridine nucleotides, based on the interconversion of 2-[U-14C]ketoglutarate or 2-keto[3,4-3H]glutarate and labelled L-glutamate in the reaction catalyzed by glutamate dehydrogenase, was applied to the measurement of lactate dehydrogenase activity in rat pancreatic islet homogenates. Using the tritiated tracer, the limit of sensitivity of the procedure for NAD(P)H assay was close to 1.0 fmol/sample, and lactate dehydrogenase activity could be measured in as little as 0.0005 islet/sample i.e., at a single cell level. This radioisotopic procedure, which can be used for the assay of various metabolites and enzymic activities, thus provides a tool for investigating the heterogeneity in metabolic behaviour of individual cells.
基于谷氨酸脱氢酶催化的反应中2-[U-¹⁴C]酮戊二酸或2-酮[3,4-³H]戊二酸与标记的L-谷氨酸的相互转化，一种用于测定还原型或氧化型吡啶核苷酸的放射性同位素方法被应用于大鼠胰岛匀浆中乳酸脱氢酶活性的测量。使用氚标记的示踪剂，该方法用于NAD(P)H测定的灵敏度极限接近1.0 fmol/样品，并且乳酸脱氢酶活性可以低至0.0005个胰岛/样品即单个细胞水平进行测量。这种可用于各种代谢物和酶活性测定的放射性同位素方法，因此为研究单个细胞代谢行为的异质性提供了一种工具。
